CysLTR1 Pathway Regulating Autophagy Participates In The Mechanism Of Mechanism Of Liver Injury Caused By Aluminum Overload

Objective:1.To observe the expressions of diverse subtypes of LTs receptors in rat liver exposed to chronic aluminum（Al）overload.2.To investigate the effect of CysLTR1 signal pathway intervention on Al-overload liver injury and autophagy pathway.Methods:1.SD rats were given intragastric administration o f Al-gluconate(Al³⁺200mg/kg)once a day,5 days per week for 20 consecutive weeks to establish the rat liver injury model.HE staining was used to observe the morphological changes of liver tissue.The ALT and AST in serum,as well as SOD activity and MDA content in liver tissue were detected by biochemical method.The contents of LTB4,LTC4,LTD4and LTE4 in rat livers were detected by ELISA assay.The mRN A levels of LTs receptor in rat liver tissue were detected by Real-Time Q uantitative PCR.And the expressions of LTs receptor,PI3K,AKT,p AKT,p-mTOR,p62,Beclin-1 and LC3B-II proteins were measured by western blot assay.2.Normal human liver L02 cells were randomly divided into groups:normal control group,Al overload model group（Al maltol;200μM,36h）,MK-571（3nM）+Al group,rapamycin+MK-571+Al group and wortmanin+MK-571+Al group.Cell viability was measured by MTT assay,LDH leakage rate was detected by the kit,and apoptosis rate was measured by flow cytometry（FCM）.The activity of ALT and AST were measured by the kit.The protein expressions of CysLTR1,PI3K,AKT,pAKT,p-mTO R,p62,Beclin-1 and LC3B-II were detected by western blot assay.The number of intracellular autolysosome was observed by electron microscopy.Results:1.The hepatocytes showed obvious vacuolization and spotty necrosis in Al-induced rat livers.The activity of ALT and AST in serum were significantly increased,the activity of SOD in liver was decreased,while the content of MDA was increased in rats treated with chronic Al overload.The content of LTC4 and LTD4increased significantly,but there was no significant change in LTB4 and LTE4.The levels of CysLTR1,CysLTR2 and LTB4R1 mRNA expression were obviously increased,besides LTB4R4 mRNA.At the same time,the expression of LTB4R1,CysLTR1,PI3K,AKT,mTOR and p62 proteins were increased significantly,and the expression of LC3B-II protein was significantly decreased,while the expression of LTB4R2 and CysLTR2 proteins did not change significantly.2.The CysLTR1 inhibitor（MK-571;3 nM,36 hours）can effectively improved the survival rate,reduced the LDH leakage rate,and decreased the rate of apoptosis in Al-induced L02 cells.The expression of PI3K,AKT,mTOR and p62 proteins were down-regulated and the LC3B-II protein level was up-regulated after treating with MK-571 in Al-induced L02 cells.Administration of wortmanin can remarkably antagonize the protective effect of MK-571 on L02 cells injury induced by Al overload,while rapamycin had synergistic action on the effect of MK-571.Conclusions:The expression of CysLTR1 in liver tissue of rats exposed to aluminum was significantly increased.The CysLTR1 inhibitor has a significantly protective effect on rat liver injury caused by Al.The hepatoprotective mechanism of CysLTR1 inhibitor may be related to alleviating oxidative stress and regulating PI3K/AKT/mTOR signaling pathway.Cys LTR1-PI3K/AKT/mTOR pathway may represent a potential target for the development of new drugs for chronic non-infectious liver injury.